A multiplex PCR tool for the specific identification of Oesophagostomum spp. from pigs

Abstract

Based on the sequences of the internal transcribed spacers (ITS-1 and ITS-2) of nuclear ribosomal deoxyribonucleic acid (DNA) of the porcine nodule worms Oesophagostomum dentatum and O. quadrispinulatum, a pair of specific primers (OdspF/OdspR2) for O. dentatum and a pair of specific primers (OqspF/OqspR) for O. quadrispinulatum were designed and used to develop a multiplex polymerase chain reaction (PCR) assay for the identification and differentiation of the two porcine nodule worms. This approach allowed the specific identification and differentiation of O. dentatum and O. quadrispinulatum, with no amplicons being amplified from heterogeneous DNA samples, and sequencing confirmed the identity of the sequences amplified. The minimum amount of DNA detectable using this multiplex PCR assay was 0.1 ng for both O. dentatum and O. quadrispinulatum. The identity of 53 porcine nodule worms collected from pigs from different geographical localities in mainland China was ascertained as O. dentatum or O. quadrispinulatum, respectively, by this multiplex PCR method. This multiplex PCR assay is useful for the simultaneous identification of eggs of O. dentatum and O. quadrispinulatum and should provide a useful tool for the diagnosis and molecular epidemiological investigation of Oesophagostomum spp. infection in pigs.